To analyze the structure of ions, to clearly identify substances, or even to examine complex substance mixtures, the method of "daughter spectra" acquisition of selected "parent ions" is frequently applied. Daughter spectra are mass spectra of charged fragments of selected parent ions from the primary spectrum of the substance or the substance mixture.
Selection of the parent ions relates to their mass-to-charge ratio, or more precisely, to their nominal mass-to-charge ratio, calculated from their nominal mass, i.e. a mass number which only takes into account the number of protons and neutrons in the molecule and not the precise isotope masses.
Consequently, parent ions are selected for the daughter spectrum which all have the same nominal mass. These ions will be referred to as parent ions, without regard whether these parent ions are of the same type, i.e. have the same total formula and the same ionic structure, or not.
In a first step during the analysis, the parent ions are isolated in the ion trap. This means that the ions of this nominal mass are kept stored and all the other types of ions are removed from the ion trap. This step of isolation is not always necessary, for example, when there are no ions of smaller masses in the trap. There exists a number of well-known methods for this isolation process for the parent ions.
In a second step, the parent ions are dissociated into partially charged and partially uncharged fragments by pumping adequate energy into the inner oscillation system of the molecule. This process is generally called fragmentation or, more specific for a special method, collisionally induced decomposition (CID). The charged fragments form the daughter ions.
In the final step, abundancies and masses of the charged fragment ions are determined by measurement. These pairs of values, abundancies and masses of the fragment ions, form the daughter ion spectrum, from which information about the ionic structure, identity or mixture of the parent ions can be obtained.
Structural analyses is of interest in many different investigations: it reveals, for instance, the brutto formula of the original substance, the functional sub-group composition of a molecule, particularly the amino acid sequence of peptides, proteins, proteoglycanes, or nucleotides; and last but not least the folding structure of large biomolecules if these biomolecules are subjected to certain surface reactions like deuterization.
Fragmentation of an ion takes place if sufficient "inner energy" is imparted on the ion, i.e. energy which is pumped into the inner structure of the molecule. For fragmentation there are two basically different methods of imparting energy on the ion:
1. Fragmentation by photon irradiation. This method is very efficient and provides good, frequently very characteristic fragmentation results; however, it calls for the use of strong light sources, preferably lasers. These light sources constitute an expensive feature which is not normally found on an ion trap. This type of fragmentation will not be dealt with here. PA1 2. Fragmentation by collisions with molecules of a collision gas (CID) in the ion trap. This collisional fragmentation is simple and requires no additional experimental equipment apart from what is already available to operate ion traps.
Collisionally induced decomposition of the parent ions begins when the secular oscillation of the ions in the storage field is excited by resonance with an RF field generated by applying an RF dipole voltage across the end caps, as described in U.S. Pat. No. 4,736,101 (Syka, Louris, Kelley, Stafford and Reynolds). The ions absorb energy in the dipole field and continuously enlarge their oscillation amplitudes. Because the ion trap usually contains a collision gas to damp the ionic movement, many collisions with the collision gas occur. The collision gas is normally controlled in such a way that one collision takes place in five to twenty ion oscillations. This corresponds to a collision gas pressure somewhere in the range between 10.sup.-4 and 10.sup.-2 millibar. With correct control of collision gas pressure and dipole voltage the oscillation amplitude can be just damped enough by the numerous collisions with the collision gas so that the ions do not hit the end caps. This is, however, a balance difficult to maintain.
The oscillating parent ions absorb several discrete portions of energy in subsequent collisions. These portions of energy are stored in the inner oscillation states of the ion. When a threshold value for the inner energy is exceeded, fragmentation can occur. The ions can therefore decompose although the energy taken up in an individual collision is not sufficiently high for fragmentation.
On the other hand, the absorbed portions of energy cannot be infinitely small. Due to the quantum structure of the energy levels of the ions' inner oscillation system, only discrete quantities of energy above a threshold can be absorbed. All collisions, the energy transfer of which would not be adequate to change the quantum level, behave like fully elastic collisions which take place without any energy transfer into the molecule. Only through the existence of this lower threshold value is it possible to store molecular ions in an ion trap for virtually any length of time without decomposition although the trap contains a collision gas which, due to the usual heating of the ion trap, is between room temperature and about 250.degree. C.
All the ion traps used as mass spectrometers nowadays deviate from pure quadrupole traps in order to achieve good levels of mass resolution for ion ejection during scanning. Usually weak higher even multipole fields (octopole field, dodecapole field, etc) are superposed on the pure quadrupole field. Superposition is caused simply by designing the shape of the electrode structure different from that of a pure quadrupole ion trap. Superposition with higher multipole fields results in field along the axis of the rotationally symmetric ion trap which increases not simply linearly from the center outward to the end caps, as with a pure quadrupole field, but increases disproportionally. An octopole field provides a field component which rises cubically and a dodecapole field provides a component which increases by the fifth power. The resulting ion traps, therefore, are called nonlinear ion traps.
The process of fragmentation, however, is thus impaired. If an ion increases its oscillation amplitude by resonance with the dipole field, the ion is now subjected to a retroactive force which no longer increases proportionally to the distance from the center. Consequently we no longer have a purely harmonic oscillation which is characterized by a frequency which is always constant irrespective of the oscillation amplitude. The retroactive force which increases more than proportional by the superimposed multipole fields, causes a change in oscillation frequency with increasing amplitude. The oscillation becomes faster with larger amplitudes.
With conventional ion traps having approximately 2% octopole field, measured as the additive field strength of the octopole field at the summit of the end caps compared with the field strength of the quadrupole field, the frequency shift is quite substancial. The frequency shift amounts to several percent if the ion oscillates just up to the end cap electrodes. Measured on the mass scale, the shift also accounts for several percent, which means several mass units for an ion of 100 to 200 atomic mass units. To express it more exactly: the frequency of an ion oscillating far up to the end caps equals the frequency of an weakly oscillating ion with a mass which differs by several mass units.
The ion to be fragmented therefore falls out of resonance with the applied RF dipole voltage when its oscillation amplitude increases. Further excitation of its oscillation is no longer possible. The fragmentation process is therefore very difficult.
EP 0 580 986 A1 proposes an improvement of the collisionally induced decomposition by modulation of the storage quadrupole RF voltage at the rate of the secular frequency. This method, however, underlies the same principles of frequency shift and is of no help here.
The usual method to overcome the problem with frequency shifts is a slightly nonresonant excitation of the basic oscillation of the parent ions on the flank of the resonance curve. This is achieved by slightly detuning the excitation frequency and increasing the excitation voltage. When the correct flank is chosen, and the oscillation amplitude starts to increase, the frequency of the ions moves by itself into the resonance maximum. If the amplitude then increases further, the frequency moves out of the resonance, the ions no longer resonate. Since the width of the resonance curve, however, is very small compared to the shift in secular oscillation frequency, this balancing act is usually quite unsatisfactory. M. Wang and G. Wells ("Non-Resonance Excitation and Ejection in Ion Traps", 41st ASMS Conf. Mass Spectrom. & Allied Topics, p.463, 1993) employ, for these reasons, a completely different method of exciting the ionic oscillation by superimposing low frequency DC pulses. However, this method has the disadvantage of not only acting on the parent ions to be fragmented but on all the ions in the ion trap, particularly including the daughter ions formed.
It is the task of the invention to find a method by which the secular oscillations of the parent ions can be excited in such a way that, irrespective of the pressure of the collision gas, they have an optimal amplitude for collisionally induced decomposition. On the other hand, they have to be retained from hitting the end caps, thereby being discharged and thus eliminated from the process.